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Background: The increasing use of nanotechnology means that nanomaterials will enter the environment. Ecotoxicological data are therefore required so that adequate risk assessments can be carried out. In this study, we used a standardized earthworm reproduction test with Eisenia andrei to evaluate three types of TiO2 nanoparticles (NM-101, NM-102, NM-103). The test was performed in natural sandy soil (RefeSol 01A) following Organisation for Economic Co-operation and Development Test Guideline No. 222. The nanoparticles differed in several aspects, such as crystalline structure, size, and the presence or absence of a coating. Results: Uncoated nanoparticles stimulated earthworm reproduction in a concentration-dependent manner during winter testing, increasing the number of offspring by up to 50% compared to the control. However, there was no stimulation when the same test was performed in the summer. This reflected an underlying circannual rhythm observed in the control soil, characterized by the production of a significantly larger number of juveniles in summer compared with that in winter. The effect of the uncoated TiO2 nanoparticles was to reduce or eliminate the circannual differences by increasing the reproductive rate in winter. Coated TiO2 nanoparticles did not influence earthworm reproduction. Conclusion: TiO2 appears to affect earthworm reproductive activity by abolishing the circannual rhythm that depresses reproduction in the winter. Further experiments will be necessary to determine (1) the mode of action of the nanoparticles, (2) the important parameters causing the effect (e.g., relevant soil parameters), and (3) the environmental relevance of continuous earthworm reproduction we observed under laboratory conditions

Effect of TiO2 nanoparticles in the earthworm reproduction test